This invention concerns the amino acid sequence, proline-histidine-serine-arginine-asparagine (SEQ ID NO: 1), which is an activity expression site of fibronectin, and a chemical substance, with which both terminals of the above amino acid sequence are modified (these shall be referred to hereinafter as PHSRN (SEQ ID NO: 1) and Ac-Pro-His-Ser-Arg-Asn-NH2). This invention also concerns either or both of an opthalmological treatment composition and a preventive composition having a salt of the above-mentioned substance that is allowable as a medical drug as an effective component thereof. This composition particularly concerns either or both of a preventive agent and a treatment agent for corneal disorder that exhibit wound healing promotion actions for corneal epithelium.
The cornea is a thin tissue of 0.52 mm to 1.0 mm thickness. The cornea is positioned at the foremost portion of the eyeball and is a highly differentiated tissue having a transmitting property and an appropriate refractive power for guiding light from the exterior to the receptors of the retina. The cornea has extremely important physiological functions. The structure of the cornea is comparatively simple. That is, the cornea has a highly ordered, microscopic, five-layer structure comprising the epithelial layer, Bowman's membrane, corneal stroma, Descemet's membrane, and endothelial cell layer.
Fibronectin is a glycoprotein with a molecular weight of approximately 440 thousand that is involved in cell adhesion and spreading and serves an important role in wound healing actions as well as in morphogenesis, development, and other biological phenomena. This fibronectin is a dimer in which two subunits of a molecular weight of 220 to 250 thousand each are bonded. Fibronectin has a domain structure. Fibronectin is involved in cell adhesion and bonds specifically with various extracellular matrices and bridges to fibronectin receptors (integrins) on cell surfaces.
Corneal disorder is induced by corneal ulcer, corneal epithelial erosion, keratitis, dry eye, and various other diseases. Such disorder is repaired naturally if there is no concurrent mixed infection. The principles of repair are: (1) the appearance of fibronectin at exposed corneal stroma portions at defective epithelial portions resulting from corneal disorder; (2) the binding of this fibronectin onto a matrix; and (3) the spreading and moving of epithelial cells onto this matrix. As the cornea is cured, fibronectin disappears from the damaged corneal portions.
Due to some reason, the repair process may be delayed or the epithelial defect may persist without being repaired. In such a case, the normal structuring of the epithelium is affected adversely and even the structures and functions of the stroma and endothelium may become impaired. Conventional treatment methods are passive methods in which the corneal surface is protected from external irritation in order to allow the epithelium to spread naturally and resurface the defective portions. With recent developments in cell biology, factors involved in the division, movement, adhesion, spreading, etc., of cells have become clarified. In regard to the repair of corneal epithelial defects, compounds that promote the spreading of the corneal epithelium have come to be emphasized.
Components, such as fibronectin, EGF (Epidermal Growth Factor), and hyaluronic acid, are known as treatment agents for corneal epithelial wounds. Fibronectin, which exists in human plasma, can be purified and used as blood product for instillation. Such an ophthalmic formulation is known to promote the resurfacing of corneal epithelial defects and the healing of epithelial wounds.
However presently, fibronectin must be purified from a patient's autologous plasma using a special purifying kit. Extreme trouble is thus taken to obtain fibronectin and a large burden is placed on the patient. Due to this reason, fibronectin is not put to adequate use even though it is clinically effective.
EGF (Epidermal Growth Factor) is a polypeptide with a molecular weight of 6000 that is known for its actions as a mitosis-promoting growth factor for corneal epithelium. It is known that when factors that inhibit the mitosis of the epithelium are present, the effects of EGF cannot be exhibited readily. In addition, in cases accompanying inflammation and in cases of diabetic keratopathy, angiogenesis occurs as a side-effect of EGF.
Hyaluronic acid is a glucosaminoglycan with a molecular weight of several million that has N-acetyl-D-glucosamine and D-glucuronic acid as constituent sugars. Hyaluronic acid is known to exhibit significant treatment effects as a treatment agent for dry eye. In regard to actions, hyaluronic acid acts on the adhesion, spreading, and movement of epithelial cells but is low in terms of epithelial cell prolification effects. Hyaluronic acid has the disadvantage of being difficult to use as an ophthalmic formulation due to increasing in viscosity at high concentration.
The peptide, PHSRN (SEQ ID NO: 1), is a pentapeptide disclosed in International Publication WO98/22617 and in “The PHSRN sequence induces extracellular matrix invasion and accelerates wound healing in obese diabetic mice,” The Journal of Clinical Investigation, 105 (11), pp. 1537-1545, 2000. In these prior-art literatures, the peptide, PHSRN (SEQ ID NO: 1), is indicated as exhibiting external wound healing effects as well as invasion and prolification suppressive effects against cancer cells. However, reports that concern the peptide, PHSRN (SEQ ID NO: 1), in relation to opthalmological fields are not known.
Satisfactory corneal disorder treatment compositions are thus not known and better compositions have been desired strongly.
As mentioned above, fibronectin is recognized to be clinically effective in opthalmological fields. However, due to problems particular to blood product (for example, sanitation problems, the large burden of having to sample a patient's autologous blood, the troublesomeness of purified fibronectin from plasma, etc.), fibronectin is not used widely. In addition, since the active site of fibronectin had not been clarified adequately, there was further room for research and development towards using fibronectin as an effective component of a corneal disorder treatment agent.
This invention has been made in view of the above circumstances and an object thereof is to find the activity expression site of fibronectin and another object thereof is to provide a composition, with which the activity expression site of fibronectin can be used as either or both of an opthalmological treatment drug and a preventive drug.